Analysis of polar urinary metabolites for metabolic phenotyping using supercritical fluid chromatography and mass spectrometry

Autor: Jeremy K. Nicholson, Ian D. Wilson, Matthew R. Lewis, Christopher Knappy, Robert S. Plumb, Norman W. Smith, Arundhuti Sen
Rok vydání: 2016
Předmět:
Formates
Clinical Biochemistry
Analytical chemistry
MODIFIERS
Acetates
Urine
01 natural sciences
Biochemistry
09 Engineering
Mass Spectrometry
Analytical Chemistry
10 Technology
Amines
Modifier
ELUTION
Chemistry
Hydrophilic interaction chromatography
Chromatography
Supercritical Fluid
General Medicine
Repeatability
6. Clean water
Physical Sciences
SEPARATION
Metabolome
Polar
Molecular Medicine
03 Chemical Sciences
Life Sciences & Biomedicine
Hydrophobic and Hydrophilic Interactions
Biochemistry & Molecular Biology
ADDITIVES
Analyte
animal structures
Resolution (mass spectrometry)
SFC
Mass spectrometry
Biochemical Research Methods
Article
DRUGS
Humans
Additive
Science & Technology
Chromatography
010405 organic chemistry
Elution
Methanol
Chemistry
Analytical
010401 analytical chemistry
Organic Chemistry
Method development
Water
STATIONARY PHASES
MS
ANALYTES
0104 chemical sciences
Ammonium Hydroxide
Supercritical fluid chromatography
Solvents
MOBILE-PHASE
Stationary phase
Chromatography
Liquid
Zdroj: Sen, A, Knappy, C, Lewis, M R, Plumb, R S, Wilson, I D, Nicholson, J K & Smith, N W 2016, ' Analysis of polar urinary metabolites for metabolic phenotyping using supercritical fluid chromatography and mass spectrometry ', Journal of Chromatography A, vol. 1449, pp. 141–155 . https://doi.org/10.1016/j.chroma.2016.04.040
Journal of Chromatography. a
ISSN: 0021-9673
DOI: 10.1016/j.chroma.2016.04.040
Popis: Highlights • An SFC method has been developed for the analysis of polar urinary metabolites. • 12 stationary phases, 9 modifier additives and 3 temperatures were evaluated. • DIOL and 2-PIC columns provide highest peak capacity and overall resolution. • Ammonium formate, ammonium hydroxide and water considerably improved separation. • Alkylamine additives should be strongly considered for polar SFC-UV analysis.
Supercritical fluid chromatography (SFC) is frequently used for the analysis and separation of non-polar metabolites, but remains relatively underutilised for the study of polar molecules, even those which pose difficulties with established reversed-phase (RP) or hydrophilic interaction liquid chromatographic (HILIC) methodologies. Here, we present a fast SFC-MS method for the analysis of medium and high-polarity (−7 ≤ cLogP ≤ 2) compounds, designed for implementation in a high-throughput metabonomics setting. Sixty polar analytes were first screened to identify those most suitable for inclusion in chromatographic test mixtures; then, a multi-dimensional method development study was conducted to determine the optimal choice of stationary phase, modifier additive and temperature for the separation of such analytes using SFC. The test mixtures were separated on a total of twelve different column chemistries at three different temperatures, using CO2-methanol-based mobile phases containing a variety of polar additives. Chromatographic performance was evaluated with a particular emphasis on peak capacity, overall resolution, peak distribution and repeatability. The results suggest that a new generation of stationary phases, specifically designed for improved robustness in mixed CO2-methanol mobile phases, can improve peak shape, peak capacity and resolution for all classes of polar analytes. A significant enhancement in chromatographic performance was observed for these urinary metabolites on the majority of the stationary phases when polar additives such as ammonium salts (formate, acetate and hydroxide) were included in the organic modifier, and the use of water or alkylamine additives was found to be beneficial for specific subsets of polar analytes. The utility of these findings was confirmed by the separation of a mixture of polar metabolites in human urine using an optimised 7 min gradient SFC method, where the use of the recommended column and co-solvent combination resulted in a significant improvement in chromatographic performance.
Databáze: OpenAIRE
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